The Paramyxoviridae are a large family of non-segmented negative-strand RNA viruses (NNV) associated with human respiratory illnesses (e.g. respiratory syncytial virus (RSV), human parainfluenza viruses) and with common childhood diseases such as measles and mumps. Owing to phylogenetic relationships, Paramyxoviridae are divided in two subfamilies, the Paramyxovirinae and the Pneumovirinae, and are classified in the order Mononegavirales with the families Rhabdoviridae, Bornaviridae and Filoviridae10. Nipah virus (NiV) is emblematic of emerging viruses; spilling over from its natural bat hosts in South East Asia, this virus causes outbreaks of respiratory and encephalic diseases in various mammals including humans′. Its mortality rate that can exceed 70% in humans, its potential for human-to-human transmission and the absence of vaccine or specific antiviral treatment classify NiV among biosafety level-4 (BSL-4) pathogens.
The genomic RNA of NiV, like that of all NNV, is condensed by a homopolymer of nucleoprotein (N), forming long helical nucleocapsids (NCs). These ribonucleoprotein complexes are the biologically active templates used for RNA synthesis by the viral RNA-dependent RNA polymerase2,3, and thus the replication of these viruses requires the continuous supply of unassembled N molecules to encapsidate the positive-sense and negative-sense progeny RNA molecules4.
Consistent with the ability of the NNV NCs to protect genomic RNA against nucleases, the N proteins comprise two globular domains, N-terminal (NNTD) and C-terminal (NCTD) that completely enwrap the RNA molecule5-8 (FIG. 1a). The N homopolymer is stabilized by lateral contacts and by the exchange of N-terminal (NTARM) and C-terminal subdomains (CTARM) between adjacent protomers5-8. The N of Paramyxovirinae has an additional long disordered C-terminal tail (NTAIL) that extends outside the NC and binds the C-terminal domain of P (PXD)11-13. The tight packaging of the RNA raises the hypothesis that N must open and close to accommodate RNA inside the binding groove upon the NC assembly process and to transiently release the RNA template upon passage of the RNA polymerase, but until now, there is no evidence of a conversion between open and closed N forms.
In the absence of other viral proteins, N has a strong tendency to polymerize and assemble on cellular RNAs. In Paramyxoviridae, but also in Rhabdoviridae and perhaps in all NNVs, a viral protein acts as a specific chaperone of nascent N and keeps it in an assembly-competent form (N0), by preventing both its polymerization and its interaction with cellular RNAs9,14. Paramyxoviridae and Rhabdoviridae P proteins are modular multifunctional proteins, which comprise a long intrinsically disordered N-terminal region (PNTR) and a C-terminal region (PCTR) with a multimerization domain (PMD) connected by a flexible linker to an NC binding domain (PXD)11,15,16 (FIG. 1a) and are therefore highly flexible in solution17. In both families, a short N-terminal region of P is sufficient to fulfill both chaperone roles9,18. Here inventors study the soluble NiV N0-P core complex to understand the mechanism of NC assembly.
When expressed in human cells, this P N0-binding domain, comprising the first 40 aa of NiV P, is able to inhibit the replication of Nipah virus. Structure-based mutagenesis validates the N0-P complex as the target for the inhibitory activity of the P peptide and for drug development against highly pathogenic members of the subfamily Paramyxovirinae including NiV. Sequence conservation in the binding interface between N and P among various members of the Paramyxovirinae subfamily raises the possibility of developing a broad spectrum drug against several viruses.